Metal ions such as copper are an essential element for human health (See, Lippard and Berg, Principles of Bioinorganic Chemistry, University Science Books, Mill Valley, Calif., which is hereby incorporated by reference). Enzymes harness the redox activity of copper to perform functions spanning energy generation, neurotransmitter and pigment synthesis, and epigenetic modification. On the other hand, misregulation of copper homeostasis is also connected to many diseases, including cancer (See, Brady et al., 2014, Nature 509, 492-496, which is hereby incorporated by reference) neurodegenerative Alzheimer's, Parkinson's, and Huntington's diseases (Que et al., 2008, Chem. Rev. 108, 1517-1549; and Barnham and Bush, 2014, Chem. Soc. Rev. 43, 6727-6749, each of which is hereby incorporated by reference), and genetic disorders such as Menkes and Wilson's diseases (Kaler, 2011, Nat. Rev. Neurol. 7, 15-29; Burkhead et al., 2011, Biometals, 24, 455-466; Merle et al., 2007, Gut 56, 115-120; Huster et al., 2006, S. Am. J. Pathol., 168, 423-434, each of which is hereby incorporated by reference). Technologies that can monitor metal ion homeostasis may therefore serve as valuable diagnostic tools for such diseases and related conditions.
In one example of a copper-mediated disorder, Wilson's disease is caused by mutation of the gene that encodes the copper transporter ATP7B protein. Mutations in this protein may lead to hyperaccumulation of copper in the liver, brain, kidney, and cornea, which can result in lipid peroxidation and corresponding liver damage as well as neurologic and psychiatric abnormalities. See, Kaler, 2011, Nat. Rev. Neurol. 7, 15-29; Burkhead et al., 2011, Biometals 24, 455-466; Merle et al., 2007, Gut 56, 115-120; Huster et al., 2006, Am. J. Pathol. 168, 423-434; Ridge et al., 2008, PLoS One 3, e1378; Boal et al., 2009 Chem. Rev. 109, 4760-4779; Brewer, “Wilson's disease A Clinician's Guide to Recognition, Diagnosis, and Management, Springer Science+Business Media New York, 2001, Print, each of which is hereby incorporated by reference. Patients suffering from Wilson's disease also exhibit high urinary copper levels (>100 mg/day, compared to 20-40 mg/day in healthy individuals) and increased serum free copper levels (>25 μg/dL, compared to 11-25 μg/dL in healthy individuals). See Kaler, 2011, Nat. Rev. Neurol. 7, 15-29; Burkhead et al., 2011, Biometals 24, 455-466; Merle et al., 2007, Gut 56, 115-120; and Huster et al., 2006, Am. J. Pathol. 168, 423-434, each of which is hereby incorporated by reference. The source of this elevated copper is not sufficiently understood, but it is thought to derive from necrosis of damaged liver cells that are cleared through the blood stream. See, Ridge et al., 2008, PLoS One 3, e1378; and Boal and Rosenzweig, 2009, Chem. Rev. 109, 4760-4779, each of which is hereby incorporated by reference.
Wilson's disease is potentially fatal, although it is readily treated if diagnosed early in its development and before extensive tissue damage has occurred. Recognizing Wilson's disease is a challenge all its own, however, owing to a lack of targeted and readily implemented diagnostic tools. Magnetic resonance imaging (MRI) and electroencephalography (EEG) are two non-invasive techniques currently used to aid in Wilson's diagnosis. However, these techniques are not specific for Wilson's disease and instead serve primarily to identify secondary characteristics. While genetic tests can offer highly accurate diagnoses, over 300 different mutations for Wilson's disease are listed in the Human Genome Organization database and only a few are fully characterized or widespread. Thus, genetic tests based on selected exons are not globally applicable. See Huster, 2010, Best Practice & Research Clinical Gastroenterology 24, 531-539; Ala et al., 2007, Lancet 369, 397-408; Ferenci, 2006, Hum. Genet. 120, 151-159; and Bandmann et al., 2015, Lancet 14, 103-113, each of which is hereby incorporated by reference.
In contrast, non-invasive tests on biofluids such as urine and blood can alternatively provide an accurate diagnosis, although these methods can require cumbersome extraction procedures that include the concentration of urine collected over 24 hours or acid digestion of serum. Expensive characterization methods such as inductively coupled plasma mass spectrometry (ICP-MS) or atomic absorption spectroscopy (AAS) are then used for direct copper detection. See, Brewer, Wilson's disease A Clinician's Guide to Recognition, Diagnosis, and Management, Springer Science+Business Media New York, 2001, Print; Huster, 2010, Best Practice & Research Clinical Gastroenterology 24, 531-539; Ala et al., 2007, Lancet 369, 397-408; Ferenci, 2006, Hum. Genet. 120, 151-159; Bandmann et al., 2015 Lancet 14, 103-113; Orena, 1986, Biochem. Biophys. Res. Commun. 139, 822-829; and Malshe, 2011, Q J Med. 104, 775-778, each of which is hereby incorporated by reference.